Virus-like particles (VLPs) represent the best-characterized and most easily tailored nanostructures for the targeting of disease. They are the largest assemblies to be routinely analyzed by x-ray crystallography, and the smallest entities to possess a genome, and thus they occupy a unique place between the worlds of chemistry and biology. Our recent efforts have focused on the 28 nm diameter icosahedral particle derived from the coat protein of bacteriophage Q?. This structure, produced recombinantly in E. coli in high yield and purity, is very stable toward heat, variable pH, organic solvents, and conditions of chemical modification. It is also recognized by no specific receptor on mammalian cells, and therefore may be imbued with desired properties by installing functional groups on the outside surface and inside compartment of the capsid. We have developed methods to genetically encode polypeptides on the exterior of Q? VLPs while at the same time packaging multiple copies of active enzymes on the inside. The outside groups serve to bind to chosen receptors for the targeting of specific cell types or to display antibodies for the same purpose. The enzymes on the inside can convert benign prodrugs to cytotoxic agents with high catalytic efficiency, while being protected from protease attack and denaturation. We will further develop and enhance the technology to produce these particles, with the goal of more precisely controlling their composition and using more functional components. We will also apply them to proof-of-concept tests of prodrug activation both in vitro and in vivo to determine the scope and limitations of this therapeutic approach. For their eventual clinical application, the consequences of the anti-particle immune response must also be determined. We will therefore conduct the first quantitative assessment of the effects of pre-immunization on the circulation lifetime and biodistribution of this type of nanoparticle, and test sophisticated methods to diminish their immunogenicity and opsonization in the context of these goals. The result of these efforts will be new capabilities in protein nanoparticle design and production, an exciting preclinical evaluation of the merits of prodrug therapy using targeted VLP-packaged enzymes, and a major contribution to the fundamental understanding of nanoparticle targeting and avoidance of opsonization and immunological pathways.